Biochemical and Biophysical Research Communications
Inhibition of transcriptional activities of AP-1 and c-Jun by a new zinc finger protein ZNF394☆
Section snippets
Materials and methods
Construction of cDNA Library of human embryo heart. The total RNA from 20-week-human embryo heart was extracted using standard methods. The RNA was pretreated with DNase I (RNase free) to eliminate DNA contamination. mRNA preparation and reverse transcription reaction were performed using a cDNA PCR Library Kit and cDNA Synthesis Kit according to manufacturer’s protocol (TaKaRa). Briefly, 5 μg mRNA was purified from 500 μg total RNA using Rapid mRNA Purification Kit (AMRESCO). Reverse
Results and discussion
In an effort to understand the role of KRAB/(Cys)2(His)2 zinc finger transcription factors in heart development and in cell signaling pathway, we isolated homologous genes of this family using PCR amplification from a cDNA library of human early embryo. Since nucleotide sequences of the krüppel-like zinc finger motif are highly conserved, it was feasible to isolate similar genes of this family by PCR amplification with degenerate oligonucleotide primers [7]. The heart cDNA library was
Acknowledgements
We are grateful to all members of the Center for Heart Development, College of Life Sciences in Hunan Normal University, for their excellent technical assistance and encouragement. This study was supported by the National Natural Science Foundation of China (Nos. 30170479, 30270722, 30270644, and 30213904), the key project of Education Ministry of China (No. 02A028), and the pre-973 project of Ministry of Science and Technology of China (No. 2002CCC00100).
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Abbreviations: MAPK, mitogen-activated protein kinase; GPCRs, G-protein coupled receptors; MAPKK, MKK or MEK, MAPK kinase; MAPKKK or MEKK, MAPKK kinase or MEK kinase; KRAB, krüppel-associated box; LER, leucine-rich region; SRF, serum response factor; AP-1, activation protein 1; DMEM, Dulbecco’s modified Eagle’s medium; CRE, cAMP response element; DAPI, 4′,6′-diamidino-2-phenylindole hydrochloride.
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These authors contributed equally to the work.